Pharmaceutical composition, preparation and uses thereof

ABSTRACT

The present invention relates to a pharmaceutical composition comprising the combination of (i) a biocompatible nanoparticle and of (ii) a pharmaceutical compound of interest, to be administered to a subject in need of such a compound of interest, wherein the nanoparticle potentiates the compound of interest efficiency. The longest dimension of the biocompatible nanoparticle is typically between about 4 and about 500 nm, and its absolute surface charge value is of at least 10 mV (|10 mV|). The invention also relates to such a composition for use for administering the compound of interest to a subject in need thereof, wherein the nanoparticle and the compound of interest are to be administered to said subject between more than 5 minutes and about 72 hours from each other.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application is a continuation of U.S. application Ser. No.14/892,271, filed Nov. 19, 2015, which is the U.S. national stageapplication of International Patent Application No. PCT/EP2014/061296,filed May 30, 2014, which claims the benefit of U.S. Provisional PatentApplication No. 61/828,794, filed May 30, 2013.

FIELD OF THE INVENTION

The invention relates to a pharmaceutical composition comprising thecombination of (i) a biocompatible nanoparticle and (ii) a compound ofinterest, to be administered to a subject in need of such a compound,wherein the nanoparticle potentiates the compound efficiency. Thelongest dimension of the biocompatible nanoparticle is typically betweenabout 4 and about 500 nm, and its absolute surface charge value is of atleast 10 mV (|10 mV|).

The invention also relates to such a composition for use foradministering the compound of interest to a subject in need thereof,wherein the nanoparticle and the compound of interest are to beadministered to said subject sequentially, typically between more than 5minutes and about 72 hours from each other.

The combined, and typically sequential, administration to the subject ofthe biocompatible nanoparticle and of the compound of interest maintainsthe pharmaceutical (i.e. therapeutic, prophylactic or diagnostic)benefit of said compound of interest for a reduced toxicity thereof forsaid subject, or increases its pharmaceutical benefit for an equivalentor reduced toxicity, when compared to the pharmaceutical benefit andtoxicity induced by said compound when administered at the standardpharmaceutical dose.

The pharmaceutical composition of the invention typically allows areduction of at least 10% of the administered compound pharmaceuticaldose when compared to the standard pharmaceutical dose of said compoundwhile maintaining the same pharmaceutical benefit for an equivalenttoxicity, preferably a reduced toxicity, for the subject, or whileincreasing the pharmaceutical benefit for an equivalent or reducedtoxicity for the subject.

BACKGROUND

In order to ensure safety and efficacy, therapeutic compounds arerequired to be selectively delivered to their target site at an optimalrate in the subject in need thereof.

Pharmacokinetics (pK) is a branch of pharmacology dedicated to thedetermination of the fate of substances administered externally to aliving organism. This determination involves steps of measuringcompound's concentrations in all major tissues over a long enough periodof time, preferably until the compound's elimination. Pharmacokineticsis necessary to efficiently describe the compound's behavior in vivo,including the mechanisms of its absorption and distribution as well asits chemical changes in the organism. The pK profile in the blood can befitted using various programs to obtain key pK parameters thatquantitatively describe how the body handles the compound. Importantparameters include maximum concentration (C_(max)), half-life (t_(1/2)),clearance, area under curve (AUC), and mean resident time (MRT), i.e.the average time during which a compound stays in an organism. When aprolonged blood circulation of the compound formulation is observed, itis usually associated with an increased t₁₁₂, a reduced clearance, anincreased AUC, and an increased MRT. pK data are often used in decidingthe optimal dose and dose regimen for maintaining the desirable bloodconcentration in order to improve therapeutics' efficiency with minimalside effects. In addition, as is well known by the skilled person, theblood concentration of a compound is correlated with both its efficacyand toxicity in most cases, typically for free drugs.

The physico-chemical properties of therapeutic as well as prophylacticcompounds have an important impact on their pharmacokinetic andmetabolic fate in the body. Therefore, selection of appropriatephysico-chemical properties is key when designing such a compound.However, since the compound is not always endogenously provided by theorganism itself and is usually externally administered, itsbiodistribution profile has to be optimized in order to fit with, andpreferably optimize, the desired pharmacological action thereof.

Several approaches have been explored to optimize the delivery of acompound to its target site. A strategy is to design a therapeuticcompound with stealth properties to prolong its blood half-life and,consequently, to enhance its accumulation to the target site. Onefavorable approach is the covalent attachment of polyethylene glycol(PEG) to the therapeutic compound that has proved to increase the invivo half-life (t_(1/2)) of the circulating compound, the level of thein vivo half-life increase varying depending partly on the nature of thecompound and on that of the coating. Also, drug carriers such asliposomes, emulsions or micelles have been developed to enhancetherapeutic efficacy of drugs by modifying their biodistribution profilein the subject's body.

However, lack of selectivity in the biodistribution of the therapeuticcompounds still remains a concern. So far, poor pharmacokinetics andhigh toxicity are important causes of failure in therapeutic compounddevelopment.

As an example, in the context of cancer treatment, intentionalinhibition of essential functions of the body in order to kill cancercells results in on-target or on-mechanism toxicity in normal cells, andclinicians have to rely on differences in dose-response and therapeuticcompound distribution between tumors and normal tissues to find apossible therapeutic window. Of note, hepatotoxicity remains a majorreason for drug withdrawal from pharmaceutical development and clinicaluse due to direct and indirect mechanisms of drug-induced cell injury inthe liver.

An approach proposed for nanoparticulate compounds such as drug carriers[Critical Reviews in Therapeutic Drug Carrier Systems 11(1):31-59 1994]is to pre-inject a decoy carrier to decrease, saturate, or eveninactivate the phagocytic capacity of the reticuloendothelial system(RES). Impairment or blockade may also be associated with decreasedplasma levels of opsonic molecules. Intravenous administration ofcertain agents, such as alkyl esters of fatty acids, dextran sulfate,salts of rare earth elements (e.g. GdCl₃), or drug carriers, eitherempty or encapsulating clodronate, prior to administration of testparticles has been demonstrated to induce moderate to dramatic reductionin Kupffer cells uptake.

For instance, the authors of “Biomimetic amplification of nanoparticlehoming to tumors” [PNAS 2007], reported the role of RES in the clearanceof their nanoparticles “CREKA-SPIO”. Initial experiments showed thatintravenous (IV) injected “CREKA-SPIO” nanoparticles did not effectivelyaccumulate in MDA-MB-435 breast cancer xenografts. In contrast, a highconcentration of particles was seen in RES tissues. By depleting RESmacrophages in the liver with liposomal clodronate, they found a 5-foldprolongation of their particle's half-life. However, clodronate agentinduces the apoptosis of macrophages from liver and spleen, and this isconsidered globally detrimental as macrophage depletion increases therisks associated with immunosuppression and infection. In a secondexperiment, the authors tested liposomes coated with chelated Ni (II) asa potential decoy particle hypothesizing that iron oxide and Ni (II)would attract similar plasma opsonins, and that Ni-liposomes couldtherefore deplete them in the systemic circulation. Indeed, intravenous(IV) injected Ni-liposomes, whether administered 5 minutes or 48 hoursbefore the injection of CREKA-SPIO nanoparticles, allow a five-foldincrease of the nanoparticles' blood half-life. However, high toxicitywas observed causing deaths among tumor mice. Plain liposomes were alsotested instead of Ni-liposomes. However, while reducing the toxicitywhen compared to said Ni-liposomes, plain liposomes were far lesseffective than them. Indeed, the blood half-life increase was only by afactor of about 2.

WO2005086639 relates to methods of administering a desired agentselectively to a target site in a subject, typically in the context ofultrasound or X-ray exposure, or in the context of magnetic resonanceimaging (MRI), as well as in the context of therapy. The aim of thedescribed method is to improve or maintain the efficiency of the agentof interest while reducing the total dose of agents concretelyadministered thanks to concomitant administration of a decoy inactivecarrier.

The described invention employs a probability-based approach. Anon-targeted inactive agent (“inactive carrier”) is co-administered(i.e. “substantially simultaneously”) with a targeted agent of interest(present in an “active composition”) exhibiting similar physicalfeatures, in order to facilitate the evasion of the RES system by thetargeted agent of interest thereby allowing an improved uptake of theagent of interest at the desired site. This approach results in a lowerexposure of patients to the agent of interest and, as a consequence, ina lower per dosage cost of said agent of interest. The activecomposition and the decoy inactive carrier are administered within fiveminutes of each other, preferably within 2 minutes of each other, oreven less. This approach relies on the presence of a large excess ofuntargeted “carrier” or “decoy” vehicles and on the probability thatthis decoy carrier in excess will compete with the targeted agent ofinterest for an uptake by the reticuloendothelial system when suppliedin the presence of vehicles that are targeted to a desired location. Thehalf-life of particles captured by RES is dose dependent, i.e. thecirculating half-life of particles increases as the dosage increases.The slower clearance associated with higher dosages is thought to favorthe maintaining of a total agents high concentration allowing a decreaseof the dose of the agent of interest which is to be administered. Inother words, an increased half-life of total agents due to a globalhigher dosage thereof should be beneficial to the targeted agents,according to the authors of WO2005086639. The requirement involved bythis approach is that the active agent and the inactive one behavesimilarly with regard to their clearance characteristics in the RES,whatever their respective compositions.

In this approach, the quasi-concomitant injection of the inactive agentand of the active one is required to increase the global amount ofagents present in the blood and consequently to prolong their bloodhalf-life. Such strategy, which expressly relies on a probability-basedapproach, necessarily requires the association of the active agent witha targeting agent in order to achieve its successful accumulation on thetarget site by conferring said active agent an advantage over theinactive one. In addition, due to the quasi-concomitant injection, aspecific design of the inactive carrier may be required depending on theintended use of the active composition.

As is apparent from the prior art, and despite of a long medical need,the improvement of compounds (including therapeutic and prophylactic aswell as diagnostic compounds) which cannot be efficiently used inpatients due to their unacceptable toxicity or to their unfavorablepharmacokinetics parameters remains a concern.

DETAILED DESCRIPTION

The present invention now allows optimization of the efficiency of acompound of interest (herein also simply identified as “the compound”)whatever its intended use in the context of therapy, prophylaxis ordiagnostic. The composition herein described which is a combination of(i) a biocompatible nanoparticle and of (ii) at least one compound ofinterest, optimizes the at least one compound of interestpharmacokinetic parameters, and, as a consequence, now renders possiblethe development of therapeutic compounds which could not have beendeveloped otherwise due for example to their unacceptable toxicity.

A typical composition of the invention (herein generally identified as“pharmaceutical composition”) is a composition comprising thecombination of (i) a biocompatible nanoparticle and (ii) at least onecompound (“the compound of interest”), wherein the longest dimension ofthe biocompatible nanoparticle is typically between about 4 nm and about500 nm, and the absolute surface charge value of the biocompatiblenanoparticle is of at least 10 mV.

A preferred object of the invention is a pharmaceutical compositioncomprising the combination of (i) a biocompatible nanoparticle and of(ii) a pharmaceutical compound of interest, wherein the longestdimension of the biocompatible nanoparticle is between about 4 nm andabout 500 nm, and the absolute surface charge value of the biocompatiblenanoparticle is of at least 10 mV mV|) for use for administering thepharmaceutical compound of interest to a subject in need thereof,wherein the nanoparticle and the compound of interest are to beadministered to a subject in need of said compound of interest betweenmore than 5 minutes and about 72 hours from each other.

The combined administration to the subject of the biocompatiblenanoparticle and of the compound of interest, through the composition ofthe invention, typically allows (maintains) the same pharmaceutical(i.e. therapeutic, prophylactic or diagnostic) benefit of the compoundwith a reduced toxicity thereof for the subject, or increases thepharmaceutical benefit of the compound with an equivalent or reducedtoxicity thereof for the subject (preferably a reduced toxicity), whencompared to pharmaceutical benefit and toxicity induced by the standardpharmaceutical dose of said compound.

The pharmaceutical composition of the invention typically allows areduction of at least 10%, preferably at least 15%, of the administeredcompound pharmaceutical (i.e. therapeutic, prophylactic or diagnostic)dose when compared to the standard pharmaceutical dose of said compound(i) while maintaining the same pharmaceutical benefit for an equivalenttoxicity, preferably a reduced toxicity, for the subject or (ii) whileincreasing the pharmaceutical benefit for an equivalent or reducedtoxicity for the subject.

As the shape of the particle can influence its “biocompatibility”,particles having a quite homogeneous shape are herein preferred. Forpharmacokinetic reasons, nanoparticles being essentially spherical,round or ovoid in shape are thus preferred. Such a shape also favors thenanoparticle interaction with or uptake by cells. Spherical or roundshape is particularly preferred.

In the spirit of the invention, the term “nanoparticle” refers to aproduct, in particular a synthetic product, with a size in the nanometerrange, typically between about 1 nm and about 500 nm, preferably betweenabout 4 nm and about 500 nm, between about 4 and about 400 nm, about 30nm and about 300 nm, about 20 nm and about 300 nm, about 10 nm and about300 nm, for example between about 4 nm and about 100 nm, for examplebetween about 10 nm, 15 nm or 20 nm and about 100 nm, or between about100 nm and about 500 nm, typically between about 100 nm and about 300nm.

The terms “size of the nanoparticle”, “largest size of the nanoparticle”and “longest size of the nanoparticle” herein typically refer to the“longest or largest dimension of the nanoparticle” or “diameter of thenanoparticle” when spheroid or ovoid in shape. Transmission ElectronMicroscopy (TEM) or Cryo-TEM can be used to measure the size of thenanoparticle. As well, Dynamic Light Scattering (DLS) can be used tomeasure the hydrodynamic diameter of nanoparticles in solution. Thesetwo methods may further be used one after another to compare sizemeasurements and confirm said size. A preferred method is DLS (Ref.International Standard IS022412 Particle Size Analysis—Dynamic LightScattering, International Organisation for Standardisation (ISO) 2008).

To be usable in the context of the invention, the absolute electrostaticsurface charge (also herein identified as “charge” or “surface charge”)of the biocompatible nanoparticle is to be higher than |10 mV| (absolutevalue). The surface charge of a nanoparticle is typically determined byzeta potential measurements in an aqueous medium for a nanoparticleconcentration between 0.2 and 10 g/L, for a pH between 6 and 8, andtypically for electrolyte concentrations in the aqueous medium between0.001 and 0.2 M, for example, 0.01 M or 0.15 M.

Typically, the biocompatible nanoparticle of the present invention hasan electronic surface charge of at least |10 mV|, i.e. below −10 mV orabove +10 mV, for example below between −12 mV or −15 mV and −20 mV orabove between +12 mV or +15 mV and +20 mV, typically below −15 mV orabove +15 mV. Preferably, the biocompatible nanoparticle of the presentinvention has an absolute electronic surface charge value (“absolutesurface charge value”) of more than 10 mV, said charge being even morepreferably a negative charge.

So long as it is charged, the nanoparticle usable in the context of theinvention can be either organic or inorganic. A mixture of organic andinorganic nanoparticles can further be used.

When organic, the nanoparticle can be a lipid-based nanoparticle(glycerolipid, phospholipid, sterol lipid, etc.), a protein-basednanoparticle also herein identified as “protein-nanoparticle” (albuminfor instance), a polymer-based nanoparticle (“polymeric nanoparticle”),a co-polymer-based nanoparticle (“co-polymeric nanoparticle”), acarbon-based nanoparticle, a virus-like nanoparticle (for example aviral vector).

The organic nanoparticle may further be a nanosphere (plainnanoparticle) or a nanocapsule (hollow nanoparticle) such as a liposome,a gel, a hydrogel, a micelle, a dendrimer, etc. A mixture of the hereindescribed organic nanoparticles can also be used.

The polymer or co-polymer can be of natural or synthetic origin.

Examples of synthetic (artificial) and natural polymers or co-polymersusable in the context of the invention to prepare organic nanoparticlescan be selected from polylactic acid (PLA), Poly(lactide-co-glycolic)acid (PLGA), Polyethyleneglycol (PEG), Polyglactin, Polylactide,Polyoxyethylene fatty acid esters, Polypropylene glycol, Polysorbate,Polyvinyl alcohol, Polyacrylamide, Polymethylmethacrylate,Polyalkylcyanoacrylate, Polylactate-co-glycolate, Poly(amido amine),Poly(ethyleneimine), alginate, cellulose and cellulose derivatives,polymers, collagen, hyaluronic acid, polyglutamic acid (PGA), actin,polysaccharide, and gelatin.

When inorganic and when its longest dimension is typically below about10 nm, for example below about 8 nm, below about 7 nm, typicallycomprised between about 7 nm and about 4 nm, for example below about 6nm, below about 5 nm or below about 4 nm, the nanoparticle may be madeof any inorganic material. The inorganic material may for examplecomprise metallic element from period 3, 4, 5, 6 of the Mendeleev'speriodic table, including the lanthanides. When the longest dimension ofthe nanoparticle is typically below about 10 nm, the nanoparticles mayassemble in larger structures. Assembling of nanoparticles in largerstructure may typically be triggered by interactions betweennanoparticles and a biocompatible polymer(s), protein(s), etc. Largerstructure may also be obtained by trapping the nanoparticles in acarrier, typically a plain carrier such as gelatin structure (alsoherein identified as “gelatin nanoparticle”) or a hollow carrier such asliposome. After in vivo administration, those larger structures mayfurther release the nanoparticles.

When inorganic and when the longest dimension of said nanoparticle istypically of at least 10 nm, typically between 10 and 500 nm, thenanoparticle may comprise at least one of or may consist of (i) one ormore divalent metallic elements selected for example from Mg, Ca, Ba andSr, (ii) one or more trivalent metallic elements selected for examplefrom Fe and Al, and (iii) one or more tetravalent metallic elementscomprising Si.

In a particular embodiment, the inorganic material of the nanoparticleis selected from (i) one or more divalent metallic elements selected forexample from Mg, Ca, Ba and Sr (ii) one or more trivalent metallicelements selected for example from Fe and Al and (iii) one or moretetravalent metallic elements comprising Si.

In a further particular embodiment, the inorganic material of thenanoparticle is selected from calcium carbonate (CaCO₃), magnesiumcarbonate (MgCO₃), magnesium hydroxide (Mg(OH)₂), iron hydroxide(Fe(OH)₂), iron oxyhydroxide (FeOOH), iron oxide (Fe₃O₄ or Fe₂O₃),aluminium oxide (Al₃O₄), aluminium hydroxide (Al(OH)₃), aluminiumoxyhydroxide (AlOOH) and silicon oxide (SiO₂).

The nanoparticles used in the herein described compositions are to bebiocompatible, i.e. compatible with living tissues. When required bytheir composition, the nanoparticles are thus to be coated with abiocompatible material to become usable. In a particular embodiment ofthe invention, the herein mentioned nanoparticle is thus covered with abiocompatible coating.

The biocompatible material can be an agent allowing interaction with abiological target. Such an agent will typically bring a positive or anegative charge on the nanoparticle's surface when the absolute chargeof the nanoparticle is of at least 10 mV.

An agent forming a positive charge on the nanoparticle's surface can befor example selected from aminopropyltriethoxisilane or polylysine. Anagent forming a negative charge on the nanoparticle surface can be forexample selected from a phosphate (for example a polyphosphate, ametaphosphate, a pyrophosphate, etc.), a carboxylate (for examplecitrate or dicarboxylic acid, in particular succinic acid) or asulphate.

In a particular embodiment, as long as the absolute charge of thenanoparticle is of at least 10 mV (|10 mV|), the nanoparticle can becoated with a biocompatible material selected from an agent displaying asteric group. Such a group may be selected for example from polyethyleneglycol (PEG); polyethylenoxide; polyvinyl alcohol; polyacrylate;polyacrylamide (poly(N-isopropylacrylamide)); polycarbamide; abiopolymer; a polysaccharide such as dextran, xylan and cellulose;collagen; a switterionic compound such as polysulfobetain; etc.

The biocompatible coating may advantageously be a “full coating”(complete monolayer). This implies the presence of a very high densityof biocompatible molecules creating an appropriate charge on the allsurface of the nanoparticle.

The biocompatible coating may further comprise a labelling agent,typically an agent allowing the visualisation of a color using standardimaging equipment.

The combined administration of the biocompatible nanoparticle togetherwith the compound of interest maintains the pharmaceutical (i.e.therapeutic, prophylactic or diagnostic), typically therapeutic, benefitof the compound of interest for a reduced toxicity, or increases thepharmaceutical benefit of the compound for an equivalent or reducedtoxicity, for the subject, typically when administered to the subject inneed of the compound of interest between more than 5 minutes and about72 hours from each other, when compared to pharmaceutical benefit andtoxicity induced by the standard pharmaceutical, typically therapeutic,dose of said compound.

In a particular embodiment, the combined administration of thebiocompatible nanoparticle and of the compound of interest allows areduction of at least 10%, preferably at least 15%, of the administeredcompound therapeutic dose, typically when administered to the subject inneed of the compound of interest, between more than 5 minutes and about72 hours from each other, when compared to the standard therapeutic doseof said compound while maintaining the same therapeutic benefit for anequivalent toxicity or a reduced toxicity (preferably a reducedtoxicity) of the compound for the subject; or while increasing thetherapeutic benefit for an equivalent or reduced toxicity of thecompound for the subject.

In a particular embodiment, nanoparticle(s) are administered withseveral compounds of interest, typically two compounds of interest.

The nanoparticle is preferably cleared from the subject to whom it hasbeen administered typically within 1 hour and 6 weeks, for example 1month (4 weeks), within 1 hour and 1 month, for example between 1 hourand 3 weeks, or between 1 hour and 2 weeks, or between 1 hour and 1week, following its administration to a subject in need of the compoundof interest.

The material constituting the nanoparticle (including its biocompatiblecoating when present) is important in determining the biopersistence ofthe nanoparticle. The nanoparticle may be regarded as biodegradable(when constituted for example by a biodegradable polymer such as PLGA orPLA), dissolvable (iron oxide for example) or non biodegradable and nondissolvable. Biodegradable and dissolvable nanoparticles facilitaterapid nanoparticle clearance from the subject.

Different molecules or agents can be used according to the presentteaching as the at least one compound of interest, typically as the atleast one pharmaceutical compound of interest, administered incombination with a biocompatible nanoparticle as described hereinabove.This compound may be a therapeutic, a prophylactic or a diagnosticcompound as previously explained. It can be an organic compound or aninorganic compound.

Examples of organic compounds usable as the compound of interest can beselected from a biological compound, an antibody, an oligonucleotide, asynthesized peptide, a small molecule targeted therapeutic, a cytotoxiccompound, and any corresponding prodrug or derivative thereof, etc.

In a particular embodiment, the compound of interest used in the contextof the present invention is an organic compound preferably selected froma biological compound, a small molecule targeted therapeutic, and acytotoxic compound. In another particular embodiment, the compound ofinterest is selected from an antibody, an oligonucleotide, and asynthesized peptide.

A biological compound is for instance an antibody, preferably amonoclonal antibody (“mAb”), such as infliximab, adalimumab,bevacizumab, rituximab, trastuzumab, ranibizumab, cetuximab, orpanatimumab; a protein or a recombinant protein, such as enbrel(etanercept) or interferon beta-la; a peptide or a recombinant peptidesuch as insulin glargine or betaseron; a vaccine such as prevnar 13 orgardasil; a biosimilar such as epogin; an enzyme or a recombinant enzymesuch as replagal or creon; etc.

An oligonucleotide is for instance an antisense oligonucleotide, anaptamer, such as mipomersen sodium or pursennid, etc.

A synthesized or artificial peptide such as glatiramer acetate orleuprolide acetate.

A small molecule targeted therapeutic generally inhibits enzymaticdomains on mutated, overexpressed, or otherwise critical protein(potential target in the context of cancer treatment) within themalignant cells. Some therapeutic agents include those that target celldivision (for example, an aurora-kinase inhibitor or a cyclin-dependentkinase inhibitor), as well as other biological mechanisms such asprotein turnover and chromatin modification (for example ahistone-deacetylase inhibitor). Small molecules targeted therapeuticsare for instance imatinib, rapamycin, gefitinib, erlotinib, sorafenib,sunitinib, nilotinib, dasatinib, lapatinib, bortezomib, atorvastatin,etc.

A cytotoxic compound is for instance a DNA-modifying agent, such as ananthracycline (for example doxorubicine, daunorubicine, etc.), analkylating agent (for example melphalan or temozolomide), as well as adrug interfering very precisely with defined physiological mechanismssuch as microtubule polymerization (for example taxol), or metabolitesynthesis (for example methotrexate). An activable cytotoxic compound istypically used in the context of Photodynamic Therapy (for examplephotofrin), and is to be activated by an external source such as a lasersource to produce its therapeutic effect. Other cytotoxic compounds aretypically selected from chemotherapeutic agents as herein described oras known by the skilled oncologist.

A prodrug (for instance capecitabine or irinotecan) is metabolized inits active form in vivo to produce its expected therapeutic effect.

Examples of inorganic compounds usable as the compound of interest canbe selected from a transition metal coordination complex, aradiopharmaceutical compound, a nanoparticle, etc.

Transition metal coordination complexes offer potential advantages overthe more common organic-based drugs, including a wide range ofcoordination numbers and geometries, accessible redox states,‘tune-ability’ of the thermodynamics and kinetics of ligandsubstitution, as well as a wide structural diversity. Metal-basedsubstances interact with cell molecular targets, affecting biochemicalfunctions resulting in malignant cell destruction. Transition metalcoordination complexes are typically cytotoxic agents (for instance,platinum coordination complexes: cisplatin, carboplatin, oxaloplatin, orruthenium or gold coordination complexes) acting on DNA structures.

Radiopharmaceutical compounds emit radiations for diagnosis purposes orin order to selectively destroy malignant cells. Typicalradiopharmaceuticals may contain for example strontium-89, thallium-201,techtenium-99, samarium-83, etc.

Nanoparticle may be selected typically from a metal oxide nanoparticle(see WO 2009/147214 and WO 2007/118884 for example), a metallicnanoparticle (gold, platinum or silver nanoparticle for instance), ametal sulfide nanoparticle (Bi₂S₃ for instance), and any mixture thereof(for example a gold nanoparticle covered with hafnium oxide material).The nanoparticle is for example a nanoparticle which can be activatedvia an external source such as an electromagnetic radiation source, aultrasound source, or a magnetic source, etc.

The compound of interest, which is administered in combination with abiocompatible nanoparticle as described hereinabove (typicallysequentially administered as herein described), may be encapsulated in acarrier or grafted (or bound) to such a carrier according to means knownby the skilled person. A typical carrier is for example a liposome (suchas DOXIL or ThermoDox which uses thermosensitive lipid), micelle,polymeric (or “polymer”) carrier, hydrogel, gel, co-polymeric carrier,protein carrier, inorganic carrier.

The pharmaceutical composition of the invention (defined by thecombination of the compound of interest and of the nanoparticle) can beused in many fields, particularly in human or veterinary medicine. Thiscomposition is typically for use in an animal, preferably in a mammal(for example in the context of veterinary medicine), even morepreferably in a human being whatever its age or sex.

The pharmaceutical compositions of the invention can be used incardiovascular diseases, Central Nervous System (CNS) diseases,gastrointestinal diseases, genetic disorders, hematological disorders,hormonal disorders, immunology, infectious diseases, metabolicdisorders, musculoskeletal disorders, oncology, respiratory diseases,toxicology, etc. In a preferred embodiment, the pharmaceuticalcomposition is used in cardiovascular diseases, CNS diseases, oncology,infectious diseases, metabolic disorders.

In the context of the present invention, the nanoparticle and thecompound(s) (“compound(s) of interest”) are advantageously to beadministered to a subject in need of said compound between more than 5minutes and about 72 hours from each other, typically between more than5 minutes and about 24 hours, preferably between more than 5 minutes or30 minutes and about 12 hours, in order to optimize the compoundpharmaceutical efficacy.

In the present invention, when the nanoparticle and the compound(s)(“compound(s) of interest”) are advantageously to be administered to asubject in need of said compound between more than 5 minutes and about72 hours from each other, the absolute surface charge value of thebiocompatible nanoparticle is of at least 10 mV mV|).

In a particular embodiment of the present invention, when thenanoparticle and the compound(s) (“compound(s) of interest”) areadvantageously to be administered to a subject in need of said compoundbetween more than 5 minutes and about 24 hours from each other, theabsolute surface charge value of the biocompatible nanoparticle isadvantageously of at least 15 mV (|15 mV|).

In another particular embodiment of the present invention, when thenanoparticle and the compound(s) (“compound(s) of interest”) areadvantageously to be administered to a subject in need of said compoundbetween more than 5 minutes and about 12 hours from each other, theabsolute surface charge value of the biocompatible nanoparticle isadvantageously of at least 20 mV (|20 mV|).

Also herein described is a method for treating a subject suffering froma disease such as those herein mentioned, wherein said method comprisesadministering to said subject a pharmaceutical composition of theinvention, typically administering a biocompatible nanoparticle and atleast one compound of interest as herein described. Any one of thenanoparticle or at least one compound of interest can be administeredfirst to the subject as long as the biocompatible nanoparticle and thecompound are administered between more than 5 minutes and about 72 hoursfrom each other. Administration of any of said nanoparticle or at leastone compound of interest can be a single administration of each,repeated administrations of each, for example several consecutiveadministrations of each. The biocompatible nanoparticle may beadministered once and the at least one compound of interest may beadministered more than once and vice versa.

In a particular embodiment, the biocompatible nanoparticle is at leastadministered at the beginning of a protocol comprising severaladministrations of a compound interest, i.e. at least at the firstadministration of said compound of interest and before or after theadministration thereof.

In another particular embodiment, the biocompatible nanoparticle is notadministered at the beginning of a protocol comprising severaladministrations of a compound interest and is not administered beforethe second or third administration of said compound of interest, andbefore or after the administration thereof.

In the context of these last two embodiments, the biocompatiblenanoparticle can also be administered together (before or after aspreviously explained) with the compound of interest during part or allof the subsequent administrations of said compound of interest.

In a particular embodiment, the nanoparticle of the invention isadministered to the subject before administration to said subject of theat least one compound of interest, typically between more than 5 minutesand about 72 hours before administration of the at least one compound ofinterest.

In this context, the term “nanoparticle” can more particularly refer toa product, in particular a synthetic product, with a size between about4 nm and about 100 nm, for example between about 10 nm, 15 nm or 20 nmand about 100 nm. An example of a compound interest to be used with suchnanoparticles is an organic compound, typically a biological compound.It is advantageously selected from an antibody, an oligonucleotide, asynthesized peptide, a small molecule targeted therapeutic, and acytotoxic compound and is preferably an antibody, a small moleculetargeted therapeutic and/or a cytotoxic compound. The term“nanoparticle” can otherwise refer to a product, in particular asynthetic product, with a size between about 100 nm and about 500 nm,typically between about 100 nm and about 300 nm. An example of acompound interest to be used with such nanoparticles is an inorganiccompound, typically selected from a metallic nanoparticle, a metal oxidenanoparticle, a metal sulfide nanoparticle and any mixture thereof orany compound of interest encapsulated in a carrier or grafted to such acarrier.

The biocompatible nanoparticle of the pharmaceutical composition of theinvention can be administered by any route, such as intravenous (IV),intra-arterial, and/or intraperitoneal. A preferred route ofadministration is the intravenous route.

The compound(s) of interest of the pharmaceutical composition of theinvention can be administered by different routes, such as subcutaneous,intravenous (IV), intradermal, intra-arterial, airway (inhalation),intraperitoneal, intramuscular and/or oral (per os) routes.

The following examples illustrate the invention without limiting itsscope.

BRIEF DESCRIPTION OF THE FIGURES

FIG. 1: Schematic view of possible routes for therapeutic compoundsremoval from blood circulation depending on the compound's size (longestdimension).

FIG. 2: Schematic representation of the treatment schedule for thepharmaceutical composition comprising (i) the biocompatiblenanoparticles of Example 3 and (ii) the Dox-NP® in MDA-MB-231-lucD3H2LNxenografts.

FIG. 3: Tumor re-growth delay of the pharmaceutical compositioncomprising the biocompatible nanoparticles of Example 3 and the Dox-NP®in MDA-MB-231-lucD3H2LN xenografts (mean RTV±SD).

EXAMPLES Example 1: Synthesis No. 1 of Liposomes as BiocompatibleNanoparticles

Liposomes are prepared using the lipidic film re-hydration method:

a) Lipids are solubilized in chloroform. Chloroform is finallyevaporated under a nitrogen flow. Re-hydration of the lipidic film withHEPES 20 mM and NaCl 140 mM at pH 7.4 is performed at 50° C., so thatthe lipidic concentration is 5 mM.

The following lipidic composition was used to prepare charged liposomes:DPPC

(DiPalmitoylPhosphatidylCholine): 86% mol; MPPC(MonoPalmitoylPhosphatidylcholine): 10% mol; DSPE-PEG(DiStearylPhosphatidylEthanolamine-[methoxy(PolyElthyleneGlycol)-2000]):4% mol.b) Freeze-thaw cycles are then performed 6 times, by successivelyplunging the sample into liquid nitrogen and into a water bath regulatedat 50° C.c) A thermobarrel extruder (LIPEX™ Extruder, Northern Lipids) was usedto calibrate the size of the liposomes under controlled temperature andpressure. In all cases, extrusion was performed at 50° C., under apressure of 10 bars.

Size distribution of the as-prepared liposomes was determined by dynamiclight scattering (DLS) using a Zetasizer NanoZS (Malvern Instruments)with a 633 nm HeNe laser at an angle of 90° C. The liposome suspensionwas diluted 100 times in HEPES 20 mM and NaCl 140 mM at pH 7.4. Liposomesize (i.e. hydrodynamic diameter) was equal to about 170 nm with apolydispersity index (PDI) equal to about 0.1.

As understandable by the skilled person, the desired surface charge wasobtained thanks to the selected lipidic composition, and its value wasconfirmed by zeta potential measurement using a Zetasizer NanoZS(Malvern Instruments).

The liposomes were diluted 100 times in water and the pH of theresulting suspension was adjusted to pH 7.4. The liposome surface chargewas equal to about −14 mV at pH 7.4.

Example 2: Method Allowing a Reduction of at Least 10% of the Dose ofTherapeutic Compound to be Administered to a Subject for an EquivalentTherapeutic Efficacy Thereof in the Subject

A pharmaceutical composition according to claim 1 comprising abiocompatible nanoparticle and an activable oxide nanoparticle foranti-cancer therapy (used as “the compound” or “pharmaceuticalcompound”) which can generate electron and/or high energy photon whenexposed to ionizing radiations such as X-rays, is administered to nudemice bearing a xenografted tumor in the following manner:

-   a) administering to each nude mouse (by intravenous injection) the    biocompatible nanoparticles;-   b) between more than 5 minutes and 72 hours following step a),    administering (by intra venous injection) the therapeutic compound    to each mouse of step a) at a lower dose (10%) when compared to the    dose currently used;-   c) measuring the therapeutic compound concentration in blood or    plasma samples of each mouse to obtain the pharmacokinetic    parameters of the therapeutic compound, said concentration being    measured once or preferably several times between 1 minute and 24    hours following the therapeutic compound administration;-   d) assessing any clinical sign of toxicity after the administration    of the pharmaceutical composition; and-   e) measuring the tumor accumulation of the therapeutic compound 24    hours after its intravenous (IV) administration.

Example 3: Synthesis No. 2 of Liposomes as Biocompatible Nanoparticles

Liposomes are prepared using the lipid film re-hydration method:

a) Lipids are solubilized in chloroform. Chloroform is finallyevaporated under a nitrogen flow. Re-hydration of the lipid film withHEPES 20 mM and NaCl 140 mM at pH 7.4 is performed at 60° C., so thatthe lipid concentration is 25 mM.

The following lipid composition was used to prepare charged liposomes:DPPC (DiPalmitoylPhosphatidylCholine) 62% mol; HSPC (HydrogenatedSoybean PhosphatidylCholine) 20% mol; CHOL (Cholesterol) 16% mol; POPS(1-Palmitoyl-2-Oleoyl Phosphatidyl Serine) 1% mol; DSPE-PEG (DiStearylPhosphatidylEthanolamine-[methoxy(PolyElthyleneGlycol)-2000]) 1%mol.

b) Freeze-thaw cycles are then performed 6 times, by successivelyplunging the sample into liquid nitrogen and into a water bath regulatedat 60° C.

c) A thermobarrel extruder (LIPEX Extruder, Northern Lipids) was used tocalibrate the size of the liposomes under controlled temperature andpressure. In all cases, extrusion was performed at 60° C., under apressure of 5 bars, with a 0.1 μm pore size polyvinylidene fluoride(PVDF) membrane.

Size distribution of the as-prepared liposomes was determined by dynamiclight scattering (DLS) using a Zetasizer NanoZS (Malvern Instruments)with a 633 nm HeNe laser at an angle of 90° C. The liposome suspensionwas diluted 100 times in HEPES 20 mM and NaCl 140 mM at pH 7.4. Liposomesize (i.e. hydrodynamic diameter) was equal to about 145 nm with apolydispersity index (PDI) equal to about 0.1.

As understandable by the skilled person, the desired surface charge wasobtained thanks to the selected lipidic composition, and its value wasconfirmed by zeta potential measurement using a Zetasizer NanoZS(Malvern Instruments).

The liposomes were diluted 100 times in a sodium chloride solution at 1mM and the pH of the resulting suspension was adjusted to pH 7.4. Theliposomes' surface charge was equal to about −25 mV at pH 7.4, NaCl 1mM.

Example 4: Tumor Re-Growth Delay of the Pharmaceutical CompositionComprising the Biocompatible Nanoparticle Suspension of Example 3 andthe Dox-NP® in MDA-MB-231-lucD3H2LN Xenografts (FIGS. 2 and 3)

This study was performed to investigate the efficacy of thepharmaceutical composition comprising (i) the biocompatible nanoparticlefrom Example 3 and (ii) Dox-NP® (Liposomal Encapsulated Doxorubicin) asthe therapeutic compound of interest, in MDA-MB-231-luc-D3H2LN tumormodel xenografted on NMRI nude mice.

The human breast adenocarcinoma MDA-MB-231-luc-D3H2LN cell line waspurchased from Caliper Life Science (Villepinte, France). The cells werecultured in Minimum Essential Medium with Earle's Balanced Salt Solution(MEM/EBSS) medium supplemented with 10% fetal bovine serum, 1%non-essential amino acids, 1% L-glutamine, and 1% sodium pyruvate(Gibco).

NMRI nude mice, 6-7 weeks (20-25 g) were ordered from Janvier Labs(France). Mice were subjected to a total body irradiation of 3Gy withthe Cesium-137 irradiation device one day before the inoculation of thecancer cells for xenograft.

MDA-MB-231-luc-D3H2LN tumors were obtained by subcutaneous injection of4×10⁶ cells in 50 μL in the lower right flank of the mouse. The tumorswere grown until reaching a volume around about 100 mm³. Tumor diameterwas measured using a digital caliper and the tumor volume in mm³ wascalculated using the formula:

${{Tumor}\mspace{14mu}{volume}\mspace{14mu}\left( {mm}^{3} \right)} = \frac{{length}\mspace{14mu}({mm}) \times ({width})^{2}\left( {mm}^{2} \right)}{2}$

Mice were randomized into separate cages and identified by a number (pawtattoo). Four groups were treated as illustrated in FIG. 2.

Group 1: Sterile Glucose 5% (Control (Vehicle) Group)

Four (4) mice were intravenously (IV) injected with a sterile glucose 5%solution on day 1, day 7 and day 14. Each time (day), two injections ofglucose 5% were performed. The first injection of glucose 5% solutionwas performed 4 hours before the second injection.

Group 2: Biocompatible Nanoparticles from Example 3 (Control Group)

Four (4) mice were intravenously (IV) injected with a sterile glucose 5%solution and the biocompatible nanoparticles from Example 3 (10 ml/kg)on day 1, day 7 and day 14. Each time (day), the injection ofbiocompatible nanoparticles from Example 3 was performed 4 hours beforeinjection of the glucose 5% solution.

Group 3: Dox-NP® (3 mg/kg Doxorubicin) (Treatment Group)

Five (5) mice were intravenously (IV) injected with a sterile glucose 5%solution and Dox-NP® (3 mg/kg doxorubicin) on day 1, day 7 and day 14.Each time (day), the injection of sterile glucose 5% solution wasperformed 4 hours before the injection of Dox-NP® (3 mg/kg doxorubicin).

Group 4: Pharmaceutical Composition, i.e. the Combination of (i) theBiocompatible Nanoparticles from Example 3 and of (ii) Dox-NP® (3 mg/kgDoxorubicin) (Treatment Group)

Five (5) mice were intravenously (IV) injected with the biocompatiblenanoparticles from Example 3 (10 ml/kg) and with the Dox-NP® (3 mg/kgdoxorubicin) on day 1, day 7 and day 14. Each time (day), the injectionof biocompatible nanoparticles from Example 3 was performed 4 hoursbefore the injection of Dox-NP® (3 mg/kg doxorubicin).

The Dox-NP® (Avanti Polar Lipids; liposomal formulation of 2 mg/mldoxorubicin HCl at pH 6.5-6.8, in 10 mM histidine buffer, with 10% w/vsucrose) was injected without additional dilution at a volume requiredto obtain 3 mg/kg of injected doxorubicin.

The biocompatible nanoparticle suspension from Example 3 was usedwithout any additional dilution.

The Dox-NP® and the biocompatible nanoparticles from Example 3 wereadministrated by intravenous injection (IV) via lateral tail vein with a100 U (0.3 ml) insulin syringe (Terumo, France).

Mice were followed up for clinical signs, body weight and tumor size.

The tumor volume was estimated from two dimensional tumor volumemeasurements with a digital caliper using the following formula:

${{Tumor}\mspace{14mu}{volume}\mspace{14mu}\left( {mm}^{3} \right)} = \frac{{length}\mspace{14mu}({mm}) \times ({width})^{2}\left( {mm}^{2} \right)}{2}$

In each group, the relative tumor volume (RTV) was expressed as Vt/V₀ratio (Vt being the tumor volume on a given day during the treatment andV₀ being the tumor volume at the beginning of the treatment).

The treatment efficacy was determined using the specific growth delay(SGD) over two doubling times (one doubling time being the amount oftime it takes for the tumor to double in volume) and the optimal percentT/C value (% T/C).

The SGD was calculated over two doubling times as follows:

${SGD} = \frac{{T\; 4d\mspace{14mu}{treated}} - {T\; 4d\mspace{14mu}{control}}}{T\; 4d\mspace{14mu}{control}}$with  T 4d  being  the  time  required  for  the  tumor  to  double  twice  in  volume  (mean  RTV  from  100  mm³  up  to  400  mm³).

The Percent T/C value (“% T/C”) was calculated by dividing the median ofthe relative tumor volume of treated groups (groups 2, 3, 4) versuscontrol group (group 1) at days 1, 3, 7, 10, 13, 15, 18, 21 and 24, andby multiplying the result of said division by 100 (see Table 2). Thelowest % T/C values obtained within 2 weeks following treatmentinjection (with or without biocompatible nanoparticles as used in thecontext of the present invention) correspond to the optimal % T/Cvalues.

FIG. 3 shows the mean relative tumor volume (mean RTV) for all groups asobtained (in the conditions previously described) after IV injectionsof:

-   -   vehicle (sterile glucose 5%) on days 1, 7 and 14 (group 1);    -   biocompatible nanoparticles from Example 3, 4 hours prior to        each vehicle (sterile glucose 5%) injection on days 1, 7 and 14        (group 2);    -   Dox-NP® (3 mg/kg doxorubicin) on days 1, 7 and 14 (group 3); or    -   biocompatible nanoparticles from Example 3, 4 hours prior to the        Dox-NP® (3 mg/kg doxorubicin) injection on days 1, 7 and 14        (group 4).

As shown in FIG. 3, a marked tumor growth inhibition is observed afterthe first injection of the pharmaceutical composition comprising thecombination of (i) the biocompatible nanoparticles from Example 3 and(ii) the Dox-NP® (3 mg/kg doxorubicin), when compared to the Dox-NP® (3mg/kg doxorubicin) alone.

The time required (expressed in days) for each tumor to double twice involume (T4d) was calculated (as a measurement of the duration of thetreatment effects). T4d for the pharmaceutical composition was estimatedto about 31 days versus about 14 days for the Dox-NP® alone (Table 1).In addition the Specific Growth Delay (SGD) estimated from the tumorsgrowth over two doubling time (starting from a mean RTV of 100 mm³ up to400 mm³) was equal to about 2 for the pharmaceutical composition versusabout 0 for the Dox-NP® alone (Table 1).

TABLE 1 Table 1: Time for the tumor to double twice in volume (T4d) andSpecific Growth Delay (SGD) estimated from the tumors growth over twodoubling times. Td4 represents the number of days to reach two doublingtimes (mean RTV from 100 mm³ up to 400 mm³). The control group is thevehicle (glucose 5%) alone (-). T4d (in days) between 100 and Groups 400mm³ (mean RTV) SGD Group 1: vehicle (control group) 11 — Group 2:Biocompatible nanoparticles 11 0 from Example 3 Group 3: Dox-NP ® alone(3 mg/Kg) 14 0 Group 4: Pharmaceutical composition 31 2 comprising (i)the biocompatible nanoparticle from Example 3 and (ii) Dox-N ® (3 mg/Kg)

Furthermore, the percent T/C (% T/C) (calculated until the day ofsacrifice of group 1) decreased faster for the pharmaceuticalcomposition than for Dox-NP® alone. This demonstrates a marked impact ofthe pharmaceutical composition. The optimal % T/C of 25 observed at day24 was indeed obtained for the pharmaceutical composition, i.e. thecombination of (i) the biocompatible nanoparticles from Example 3 and(ii) the Dox-NP® (3 mg/kg doxorubicin), whereas the optimal % T/C of 38observed at day 21 was obtained for the group Dox-NP® alone (Table 2).

TABLE 2 Table 2: percent T/C (% T/C) is calculated by dividing themedian of the relative tumor volume of treated groups (groups 2, 3, 4)versus control group (group 1) at days 1, 3, 7, 10, 13, 15, 18, 21 and24, and by multiplying the result of said division by 100. Control groupis group 1 (vehicle sterile glucose 5% alone). % T/C is calculated untilday 24 which corresponds to the day of sacrifice of group 1 (controlgroup). Optimal % T/C is indicated for each group as **. Group 4:Pharmaceutical composition comprising (i) the Group 2: biocompatibleGroup 3: Dox-NP ® biocompatible nanoparticle and (ii) Days nanoparticlesalone alone (3 mg/kg) Dox-NP ® (3 mg/Kg) 1 100 100 100 3 104 126 121 7 90 106  80 10    87 **  76  60 13 103  80  55 15  98  74  45 18  98  56 43 21  87    38 **  33 24  98  40    25 **

Overall, those results showed an advantageous tumor growth delay whenusing the pharmaceutical composition of the present invention[corresponding to the combination of (i) the biocompatible nanoparticlesfrom Example 3 and of (ii) the Dox-NP® (3 mg/kg doxorubicin)], which isnot observed when the Dox-NP® (3 mg/kg doxorubicin) is used alone (i.e.in the absence of the biocompatible nanoparticles used in the context ofthe present invention). This tumor growth delay was observed when thebiocompatible nanoparticles from Example 3 and the compound of interest(the Dox-NP®) were administered sequentially, the biocompatiblenanoparticle being administered to the subjects 4 hours before theDox-NP®.

Inventors are reproducing this experiment to confirm that the sameresult is observed so long as the compound of interest and thebiocompatible nanoparticles are administered to the subject between morethan 5 minutes and about 72 hours from each other.

We claim:
 1. A method comprising a step of intravenously administering apharmaceutical compound of interest to a subject in need thereof and adistinct step of intravenously administering a biocompatiblenanoparticle to said subject, wherein the longest dimension of thebiocompatible nanoparticle is between about 4 nm and about 500 nm, andthe surface charge value of the biocompatible nanoparticle is a negativecharge below −10 mV, said nanoparticle being intravenously administeredto the subject between more than 5 minutes and about 72 hours before thepharmaceutical compound of interest, and the biocompatible nanoparticleis an organic nanoparticle free of an additional therapeutic,prophylactic or diagnostic agent, and wherein the combined intravenousadministration of the biocompatible nanoparticle and the compound ofinterest maintains the therapeutic benefit of the compound of interestwith reduced toxicity, or increases the therapeutic benefit of thecompound of interest with equivalent or reduced toxicity for the subjectwhen compared to therapeutic benefit and toxicity induced by thestandard therapeutic dose of said compound of interest.
 2. The methodaccording to claim 1, wherein the nanoparticle is selected from alipid-based nanoparticle, a protein-based nanoparticle, a polymer-basednanoparticle, a copolymer-based nanoparticle, a carbon-basednanoparticle, and a virus-like nanoparticle.
 3. The method according toclaim 1, wherein the nanoparticle is further covered with abiocompatible coating.
 4. The method according to claim 1, wherein thecombined intravenous administration of the biocompatible nanoparticleand the compound of interest allows for a reduction of at least 10% ofthe administered compound therapeutic dose when compared to the standardtherapeutic dose of said compound of interest while maintaining the sametherapeutic benefit with equivalent or reduced toxicity for the subject,or while increasing the therapeutic benefit with equivalent or reducedtoxicity for the subject.
 5. The method according to claim 1, whereinthe nanoparticle is cleared from the subject to whom it has beenadministered within one hour and six weeks after its administration. 6.The method according to claim 1, wherein the compound of interest is anorganic compound.
 7. The method according to claim 6, wherein saidorganic compound is a biological compound, a small-molecule targetedtherapeutic, or a cytotoxic compound.
 8. The method according to claim6, wherein the compound of interest is selected from an antibody, anoligonucleotide, and a synthesized peptide.
 9. The method according toclaim 1, wherein the compound of interest is an inorganic compoundselected from a metallic nanoparticle, a metal oxide nanoparticle, ametal sulfide nanoparticle or any mixture thereof.
 10. The methodaccording to claim 1, wherein the compound of interest is encapsulatedin a carrier.
 11. The method according to claim 1, wherein the compoundof interest is bound to a carrier.
 12. The method according to claim 1,wherein the nanoparticle is administered to the subject between morethan 5 minutes and about 24 hours before the pharmaceutical compound ofinterest.
 13. The method according to claim 1, wherein the nanoparticlehas a surface charge value below −12 mV.
 14. The method according toclaim 1, wherein the nanoparticle has a surface charge value below −15mV.